Consuelo Esteve

Aeromonas encheleia sp. nov., isolated from European eels

Four strains isolated from European eels in Valencia, Spain, were found to constitute a DNA relatedness group which is 0 to 50% related to the 13 species and DNA group 11 of the genus Aeromonas. Phenotypically, these strains have all of the properties that define the genus Aeromonas. However, they differ from the previously described Aeromonas species by three or more properties. The strains are positive for motility, growth at 37 degrees C, indole production, and arginine dihydrolase activity. They exhibit negative reactions in tests for growth at 42 degrees C and in thiosulfate-citrate-bile salts-sucrose medium (Oxoid), Simmons citrate tests, and tests for lysine and ornithine decarboxylase activities. They produce acid from salicin but not from L-arabinose, D-cellobiose, or lactose. All four strains hydrolyze esculin and arbutin but not elastin. They use L-serine as a sole carbon and energy source but cannot utilize L-arabinose, L-arginine, D-gluconate, or L-glutamine. The strains are resistant to ampicillin. The guanine-plus-cytosine content of the DNA is 59.4 to 60.8 mol%. The name Aeromonas encheleia sp. nov. is proposed for these strains; strain S181 (= CECT 4342) is the type strain. This new species is generally not pathogenic for eels or mice.

Occurrence of Drug-Resistant Bacteria in Two European Eel Farms

ABSTRACT The occurrence of strains that are resistant to oxolinic acid, oxytetracycline, sulfamethoxazole-trimethoprim, and nitrofurantoin among heterotrophic bacteria, including human and fish pathogens, in two freshwater eel farms was investigated. High levels of individual- and multiple-drug-resistant bacteria were detected, although sampling events were not correlated with clinical outbreaks and drug therapy.

Vibrio Species in Seawater and Mussels: Abundance and Numerical Taxonomy

Summary Qualitative and quantitative studies were performed on Vibrio species from seawater and mussel samples at a hatchery in Valencia harbour, and from market mussels. Vibrio alginolyticus and V. harveyi were the most abundant species in all samples. Other species identified were V. pelagius, V. mediterranei, V. tubiashii, V. damsella, V. splendidus, V. fluvialis, V. parahaemolyticus, V. anguillarum, V. cholerae and Aeromonas spp . The abundance of V. harveyi and V. mediterranei was positively correlated with temperature whereas V. pelagius counts showed negative correlation with this parameter. The pathogenic species were recovered in very low numbers. A numerical taxonomic study was performed on several representative isolates. Separation between V. parahaemolyticus and V. alginolyticus was not possible at species level. Two phena were phenotypically different from any described Vibrio species.

Multiresistant waterborne pathogens isolated from water reservoirs and cooling systems

Aims:  To determine the incidence of multiple antibiotic‐resistant strains of the emergent human pathogens Legionella pneumophila, Pseudomonas aeruginosa and mesophilic Aeromonas species among those isolated from water reservoirs and industrial cooling systems.

Numerical Taxonomy of Aeromonadaceae and Vibrionaceae associated with Reared Fish and Surrounding Fresh and Brackish Water

Summary A numerical taxonomic study was performed on 51 isolates of gram-negative, oxidase-positive, rodshaped, heterotrophic, facultative anaerobic bacteria isolated from aquatic samples (fresh and brackish water, eels, turbot and rainbow trout). These bacteria were compared with 28 type and references strains which included most of Aeromonas phenospecies and DNA hybridization groups, as well as strains of some arginine-positive vibrios and Plesiomonas shigelloides . The strains were characterized by 137 morphological, biochemical, physiological and nutritional tests, and data were examined by simple matching (S SM ) and Jaccard (S J ) coefficients. Unweigthed pair group mathematical averaging cluster analysis (UPGAIA) resolved 17 groups of operational taxonomic units at Ss,,, values of ≤ 86%. A total of 16 phena were defined. Larger phena were identified as Aeromonas jandaei and A. hydrophila . The species A. caviae, A. encheleia, A. allosaccharophila, A. salmonicida, A. sobria, A. veroi biovar veronii, A. media, Vibrio anguillarum, V. fumissii and Plesiomonas shigelloides were also represented. One phenon was identified as a member of the genus Aeromonas . Similar groupings were obtained at S values of ≤ 79%. Virulence trials showed that most of the strains included in phena identified as A. jandaei, A. hydrophila and V. fumissii were pathogenic for eels (sample mean LD 50 of 6.5 to 7.6 (log 10 )), whereas other aquatic isolates as well as reference strains- from clinical origin were avirulent for fish.

Mechanisms of quinolone resistance in Aeromonas species isolated from humans, water and eels

Mechanisms of resistance were determined in 33 quinolone-resistant isolates of the species Aeromonas hydrophila, Aeromonas caviae, Aeromonas media, Aeromonas salmonicida, Aeromonas popoffii and Aeromonas veronii, recovered from humans, freshwater and eels. The quinolone resistance-determining regions (QRDRs) of gyrA and parC genes were sequenced in these resistant strains, as well as in 8 quinolone-sensitive Aeromonas used as controls. All quinolone-resistant Aeromonas carried point mutations in the gyrA QRDR at codon 83, respectively giving rise to substitutions Ser(83)-->Ile (32 strains) or Ser(83)-->Val (1 strain). Almost half of these isolates (48%) carried additional point mutations in the gyrA QRDR at codon 92 and/or in the parC QRDR at codon 80 corresponding to substitutions Leu(92)-->Met and Ser(80)-->Ile. In all cases, MICs of quinolones were determined in the presence and absence of the efflux pump inhibitor phenylalanine-arginine beta-naphthylamide (PAbetaN). Addition of PAbetaN had no effect on the levels of resistance observed in these isolates. In conclusion, the mechanism of quinolone resistance in the Aeromonas isolates studied was related to mutations in QRDR regions of gyrA and parC genes, with little obvious involvement of pumps inhibited by PAbetaN.

Phenotypic study by numerical taxonomy of strains belonging to the genus Aeromonas

Aims:  This study was undertaken to cluster and identify a large collection of Aeromonas strains.

Plasmid-Mediated QnrS2 Determinant from a Clinical Aeromonas veronii Isolate

The main objective of this study was to determine the prevalence of the Qnr determinants in clinical and environmental Aeromonas spp. A total of 52 Aeromonas sp. isolates identified by biochemical methods (5), 25 isolated from natural waters (1) and 27 isolated from clinical samples from hospitals in Valencia, Spain, were tested for quinolone resistance by the disk diffusion method (4) (nalidixic acid, 30 μg; oxolinic acid, 2 μg; flumequine, 30 μg; ciprofloxacin, 5 μg; and levofloxacin, 5 μg). Among the studied isolates, 27 showed resistance to nalidixic acid and susceptibility to ciprofloxacin, 24 isolates were susceptible to both nalidixic acid and ciprofloxacin, and only 1, the A. veronii A272 clinical isolate, was resistant to both nalidixic acid and ciprofloxacin. The isolates resistant to nalidixic acid were also resistant to oxolinic acid and flumequine. Moreover, A. veronii A272 was the only one resistant to levofloxacin. Screening of the qnrA, qnrB, and qnrS genes was performed by multiplex PCR using a set of specific primers for all isolates. Bacterial strains positive for each qnr gene were used as positive controls (Klebsiella pneumoniae UAB1 for qnrA, Escherichia coli J53 pMG252 for qnrB, and E. coli J53 pMG298 for qnrS) and were run in each batch of tested samples. Only an A. veronii clinical isolate (A. veronii A272) presented a qnr gene, which showed 100% homology with the qnrS2 gene previously reported in an isolate from the bacterial community of a wastewater treatment plant in Germany (2) and in a non-Typhi Salmonella clinical isolate in the United States (6).

Pathogenic Aeromonas hydrophila Serogroup O:14 and O:81 Strains with an S Layer

ABSTRACT Five autoagglutinating Aeromonas hydrophila isolates recovered from eels and humans were assigned to serogroups O:14 and O:81 of the Sakazaki and Shimada (National Institutes of Health) scheme. They had the following properties in common: positive precipitation after boiling, moderate surface hydrophobicity (salt-aggregation-test value around 1.2), pathogenicity for fish and mice (50% lethal dose, 104.61 to 107.11), lipopolysaccharides that contained O-polysaccharide chains of homogeneous chain length, and an external S layer peripheral to the cell wall observed by electron microscopy. A strong cross-reactivity was detected by immunoblotting between the homogeneous O-polysaccharide fraction of O:14 and O:81 strains but not between them and the lipopolysaccharide of A. hydrophila TF7 (O:11 reference strain). Outer membrane fractions of these strains contained a predominant 53- to 54-kDa protein which was glycine extractable under low-pH (pH 2.8) conditions and was identified as the surface array protein. The S-layer proteins of the O:14 and O:81 A. hydrophila strains seemed to be primarily different from those previously purified from strains A. hydrophila TF7 and Aeromonas salmonicida A450 on the basis of colony hybridizations with both the structural genes vapA and ahsA. This is the first report of the presence of an S layer in mesophilic Aeromonas strains not belonging to serogroup O:11.

DNA Relatedness among Aeromonas allosaccharophila Strains and DNA Hybridization Groups of the Genus Aeromonas

The genomic relatedness among three Aeromonas allosaccharophila strains, including the type strain, and other Aeromonas type and reference strains that were assigned to DNA hybridization groups was estimated by DNA-DNA hybridization (competition procedure using a membrane method). All A. allosaccharophila strains were highly related (70 to 100%) to strains 289T (= CECT 4199T) and ATCC 35942. Type strains of other validated Aeromonas species, reference strains of DNA groups 8 and 11, and the Aeromonas sp. strain ATCC 43946 (enteric group 501) were 0 to 41% related to A. allosaccharophila 289T and ATCC 35942. The G+Cs content of A. allosaccharophila strains were in the range 55.9 to 57.3 mol%. The G+C content of the type strain of this species was 56.9 mol%, a value somewhat lower than that reported in the original description.